Complete genome sequences enable new approaches for antigen discovery. We propose to test a novel approach that uses bacterial genomes to identify candidate vaccine antigens that are up-regulated when bacteria are grown under conditions that mimic human infection. Our hypothesis is that genes that are up- regulated will include novel, surface-exposed antigens that may have gone unrecognized by previous studies of bacteria grown in artificial media. To test our hypothesis, we will use Neisseria meningitidis group B (NmB), an important cause of bacterial meningitis and sepsis, for which there is no broadly protective vaccine available. NmB is ideal for this purpose since in nature it exclusively infects humans;its hallmark is rapid replication in blood;and there exists a reliable serologic surrogate for predicting protective immunity, serum bactericidal activity. In Aim 1, we will perform transcriptional profiling using a combination of DNA microarrays and quantitative, reverse-transcription PCR to identify genes that are up-regulated when bacteria are grown in human blood or plasma. In Aim 2, we will identify genes that are up-regulated in bacteria isolated from infant rats with bacteremia. In Aim 3 we will express recombinant proteins encoded by NmB genes that are up- regulated during infection and, based on genomic data, are predicted to be conserved and surface-exposed. Mice will be immunized and the resulting antisera assayed for complement-mediated bactericidal activity against a panel of genetically diverse group B strains. Selected antisera will be tested for passive protection in the human blood ex vivo infection model and/or infant rat bacteremia model. The results may identify a new promising NmB vaccine candidate as well as determine whether similar transcription profiling studies have the potential for identifying vaccine antigens for other bloodborne pathogens. Project Narrative / Significance: We propose to evaluate a novel transcriptional profiling approach for discovery of vaccine antigens against bloodborne pathogens. We will employ Neisseria meningitidis group B, which is an ideal model organism to test our approach since it causes sepsis and meningitis exclusively in humans, no broadly protective vaccine is available and there exists a reliable serologic surrogate for predicting protective immunity. Our results may identify a new promising vaccine candidate for prevention of meningococcal disease, an important public health problem, as well as determine whether it is worth testing a similar approach for antigen discovery against other pathogens.